常规细胞遗传学和荧光原位杂交方法检测14q32/IgH基因重排

  目的 比较常规细胞遗传学（CC）,间期荧光原位杂交（FISH）技术及连续R显带后FISH检测免疫球蛋白重链（IgH）基因重排的应用价值。方法 应用常规细胞遗传学及间期FISH分析血液系统肿瘤患者43例。结果 43例患者中14q32+／IgH+患者19例,14q32-／IgH+患者2例（4.7 %）,14q32+／IgH-患者3例（7.0 %）,14q32-／IgH-患者19例。部分病例CC与间期FISH方法检测14q32／IgH基因重排得到了不一致的结果。对5例患者进行连续R显带后FISH分析,对照核型和FISH图,能清楚地看到IgH基因易位涉及的染色体。结论 间期FISH可以提高14q32／IgH重排检出率,R显带后FISH可以帮助识别与14q32易位的伙伴染色体。     

Clinical heterogeneity in childhood acute lymphoblastic leukemia with 11q23 rearrangements.

To assess the clinical heterogeneity among patients with acute lymphoblastic leukemia (ALL) and various 11q23 abnormalities, we analyzed data on 497 infants, children and young adults treated between 1983 and 1995 by 11 cooperative groups and single institutions. The substantial sample size allowed separate analyses according to age younger or older than 12 months for the various cytogenetic subsets. Infants with t(4;11) ALL had an especially dismal prognosis when their disease was characterized by a poor early response to prednisone (P=0.0005 for overall comparison; 5-year event-free survival (EFS), 0 vs 23+/-+/-12% s.e. for those with good response), or age less than 3 months (P=0.0003, 5-year EFS, 5+/-+/-5% vs 23.4+/-+/-4% for those over 3 months). A poor prednisone response also appeared to confer a worse outcome for older children with t(4;11) ALL. Hematopoietic stem cell transplantation failed to improve outcome in either age group. Among patients with t(11;19) ALL, those with a T-lineage immunophenotype, who were all over 1 year of age, had a better outcome than patients over 1 year of age with B-lineage ALL (overall comparison, P=0.065; 5-year EFS, 88+/-+/-13 vs 46+/-14%). In the heterogeneous subgroup with del(11)(q23), National Cancer Institute-Rome risk criteria based on age and leukocyte count had prognostic significance (P=0.04 for overall comparison; 5-year EFS, 64+/-+/-8% (high risk) vs 83+/-+/-6% (standard risk)). This study illustrates the marked clinical heterogeneity among and within subgroups of infants or older children with ALL and specific 11q23 abnormalities, and identifies patients at particularly high risk of failure who may benefit from innovative therapy.     

No prognostic effect of additional chromosomal abnormalities in children with acute lymphoblastic leukemia and 11q23 abnormalities.

This study characterized the additional chromosomal abnormalities (ACA) associated with 11q23 rearrangements in 450 infants and children with acute lymphoblastic leukemia (ALL) and examined the impact of these ACA on survival. Overall, 213 (47%) cases had ACA but the incidence varied according to patient age and 11q23 subgroup. Infants and patients with t(4;11)(q21;q23) had the lowest incidence of ACA (50/182 (27%) and 57/216 (26%) respectively), whereas patients with del(11)(q23) had the highest incidence (66/93 (71%)). Del(11)(q23) abnormalities were heterogeneous and occasionally secondary to t(9;22)(q34;q11.2). Thus, patients with del(11)(q23) comprised a separate biological entity, which was clearly distinct from those with an 11q23 translocation. The most frequent specific ACA were trisomy X (n = 38), abnormal 12p (n = 32), abnormal 9p (n = 28) and del(6q) (n = 19). The presence of ACA did not change the 5 year event-free survival estimates among children (56% (95% Cl 46-65%) vs 62% (54-69%)) or infants (22% (15-29%) vs 18% (9-29%)), nor when the different 11q23 subgroups were analyzed separately. This study has conclusively demonstrated that there is no prognostic effect of secondary chromosomal changes in association with 11q23 abnormalities in childhood ALL. However, characterization of these ACA is important to determine their potential role in initiation of MLL driven leukemogenesis.     

Clinical features and outcome of MLL gene rearranged acute lymphoblastic leukemia in infants with additional chromosomal abnormalities other than 11q23 translocation

The treatment outcome for infant acute lymphoblastic leukemia (ALL) with positive MLL gene rearrangements remains poor. We analyzed whether additional chromosomal abnormalities (ACA) other than 11q23 translocation could affect the disease behavior and its prognosis. Eighteen of seventy-four patients with infant acute lymphoblastic leukemia showed ACA, including three-way translocations in four, other novel translocations in four, and complex structural chromosomal changes in four. Only age less than 6 months and positive central nervous system leukemia were significant prognostic factors by multivariate analysis. However, overall survival rates were worse in patients with ACA compared to those with non-ACA. Genetic alterations induced by additional chromosomal changes may be associated with disease progression and poorer overall survival rates in infants with MLL-rearranged ALL.     

Chromosome abnormalities and MLL rearrangements in acute myeloid leukemia of infants.

Of 29 infants with acute myeloid leukemia (AML), 14 (48%) had various 11q23 translocations. MLL rearrangements were examined in 21 of the 29 patients, and 11 (52%) showed the rearrangements. 11q23 translocations and/or MLL rearrangements were found in 17 (58%) of the 29 patients. While all but one of the 17 patients with 11q23/MLL rearrangements had M4 or M5 type of the FAB classification, the 12 patients without such rearrangements had various FAB types, including M2, M4, M4EO, M6 and M7. Of the 12 patients with other chromosome abnormalities or normal karyotypes, two had inv(16) ort(16;16), one had t(1;22)(p13;q13), and two had a novel translocation, t(7;12)(q32;p13). The breakpoint on 12p of the t(7;12) was assigned to intron 1 or the region just upstream of exon 1 of the TEL/ETV6 gene by fluorescence in situ hybridization. The event-free survival at 5 years for the 17 patients with 11q23/MLL rearrangements was 42.2%, and that for the 12 patients without such rearrangements was 31.3% (P = 0.5544). 11q231MLL rearrangements have been frequently reported and a poor prognosis in infant acute lymphoblastic leukemia implied. Our study showed that while 11q23/MLL rearrangements were also common in infant AML, AML infants with such rearrangements had a clinical outcome similar to that of AML infants without such rearrangements.     

High frequency of pro-B acute lymphoblastic leukemia in adults with secondary leukemia with 11q23 abnormalities.

To evaluate the frequency and cytogenetic and immunophenotypic features of therapy-related, precursor B-cell acute lymphoblastic leukemia (ALL), 152 cases of immature B-cell ALL were reviewed. These were compared to the frequency of therapy-related acute myeloid leukemia (t-AML) during the same time period. Eight ALL cases with a prior diagnosis of malignancy were identified, including six (4.0%) with prior therapy considered to be therapy-related ALL (t-ALL). The t-ALL cases followed treatment for breast carcinoma (two cases), lung carcinoma (two cases), lymphocyte predominance Hodgkin's disease and follicular lymphoma with a latency period of 13 months to 8 years. All t-ALL cases had a pro-B (CD10-negative) immunophenotype with significantly higher expression of CD15 and CD65, compared to the de novo CD10-positive ALL cases. All six t-ALL cases had MLL abnormalities by fluorescence in situ hybridization, and four showed t(4;11)(q21;q23). These represented half of all 11q23-positive adult ALL cases. During the same time period, 4.9% of all AML cases were considered t-AML. There was a 16.7% frequency of 11q23 abnormalities in the t-AML group. Despite the similar frequency in therapy-related disease among ALL and AML cases, there were differences in the frequency of the diseases and t-ALL represented 12% of all therapy-related leukemias. However, t-ALL represented 46% of all 11q23-positive therapy-related leukemias. The immunogenetic features of t-ALL appear distinct and may aid in identifying more cases of this disease type in the future.     

Secondary acute myelogenous leukemia and myelodysplasia without abnormalities of chromosome 11q23 following treatment of acute leukemia with topoisomerase II-based chemotherapy.

Therapy-related MDS and AML are complications of intensive chemotherapy regimens. Traditionally, patients exposed to topoisomerase II inhibitors are reported to develop secondary AML with abnormalities of chromosome 11q23. We evaluated the long-term hematologic toxicity of topoisomerase II-intensive high-dose mitoxantrone-based chemotherapy in 163 newly diagnosed acute leukemia patients treated over an 8 year period. Nine (5.5%) patients developed new cytogenetic abnormalities. Four patients developed MDS with progression to AML, three patients developed new abnormalities at the time of relapse, and three patients (including one of the former patients) had changes that were not associated with hematologic disease. The abnormalities most frequently involved chromosomes 7q, 20q, 1q, and 13q. Despite the use of topoisomerase II-intensive treatment, no patient developed an abnormality involving chromosome 11q23. Spontaneous resolution of some changes and prolonged persistence of others in the absence of hematologic disease indicates that some cytogenetic changes are not sufficient to promote leukemogenesis.     

急性淋巴细胞白血病AML1基因重排与MLL基因丢失同时出现临床意义的探讨

目的：对急性淋巴细胞白血病（ALL）急性髓系白血病（AML1）基因重排与混合谱系白血病（MLL）基因丢失同时出现进行探讨。方法：在常规细胞遗传学（CC）分析基础上运用荧光原位杂交技术（FISH），采用多种位点特异性DNA探针（染色体全染、特殊位点和双色易位融合探针），对63例ALL患者（8例成人，55例儿童）进行分析。结果：63例ALL患者中有4例（6．3％）出现MLL基因重排，其中3例出现MLL基因丢失，1例出现MLL基因的移位即t（4；11），3例出现了MLL基因丢失的患者同时合并有AML1基因重排，2例为t（12；21）易位而形成的TEL／AML1融合基因，1例为AML1基因复制引起的环形21号染色体，即r（21）。55例儿童ALL中有15例（27．3％）出现AML1基因重排，即由t（12；21）易位而形成的TEL／AML1融合基因，TEL／AML1阳性患者免疫分型均为B细胞型，8例成人均无t（12；21）。结论：儿童ALL常合并有t（12；21），TEL／AML1融合基因的出现是预后良好的指标，而MLL基因重排的患者具有对常规化疗不敏感及预后不良的特点，两者同时出现说明白血病染色体重排、病理过程及影响预后因素的复杂性。     

Allogeneic stem cell transplantation for relapsed and refractory acute myeloid leukemia patients with 11q23 abnormalities

Abnormalities involving chromosome band 11q23 are seen in de novo and therapy-related acute myeloid leukemia (AML). The role of hematopoietic stem cell transplantation (SCT) in AML with 11q23 abnormalities is not well defined. We present here the outcome of 14 AML patients with 11q23 abnormalities transplanted beyond first complete remission (CR) or with primarily refractory disease. Eleven cases were de novo and three therapy-related AML. At transplant, five patients were in first untreated relapse, one second CR, one second relapse and seven had refractory disease. All 14 patients underwent allogeneic SCT. Total body irradiation was used in 93% of patients and cyclosporine-methotrexate for graft-versus-host disease prophylaxis in 71%. The relapse rate of engrafted patients was 58%. Five year survival and disease-free survival were 14 and 7%, respectively. Allogeneic SCT for AML with 11q23 abnormalities was of limited benefit in this cohort of patients transplanted beyond first CR or with primarily refractory disease.     

急性淋巴细胞白血病AML1基因重排与MLL基因丢失同时出现临床意义的探讨

目的:对急性淋巴细胞白血病(ALL)急性髓系白血病(AML1)基因重排与混合谱系白血病(MLL)基因丢失同时出现进行探讨。方法:在常规细胞遗传学(CC)分析基础上运用荧光原位杂交技术(FISH),采用多种位点特异性DNA探针(染色体全染、特殊位点和双色易位融合探针),对63例ALL患者(8例成人,55例儿童)进行分析。结果:63例ALL患者中有4例(6.3%)出现MLL基因重排,其中3例出现MLL基因丢失,1例出现MLL基因的移位即t(4;11),3例出现了MLL基因丢失的患者同时合并有AML1基因重排,2例为t(12;21)易位而形成的TEL/AML1融合基因,1例为AML1基因复制引起的环形21号染色体,即r(21)。55例儿童ALL中有15例(27.3%)出现AML1基因重排,即由t(12;21)易位而形成的TEL/AML1融合基因,TEL/AML1阳性患者免疫分型均为B细胞型,8例成人均无t(12;21)。结论:儿童ALL常合并有t(12;21),TEL/AML1融合基因的出现是预后良好的指标,而MLL基因重排的患者具有对常规化疗不敏感及预后不良的特点,两者同时出现说明白血病染色体重排、病理过程及影响预后因素的复杂性。     

The human LASP1 gene is fused to MLL in an acute myeloid leukemia with t(11;17)(q23;q21)

The MLL gene at chromosome 11q23 is frequently rearranged in acute leukemia. Here we report the identification of a new MLL fusion partner in the case of an infant with AML-M4 and a t(11;17)(q23;q21) translocation. Fluorescence in situ hybridization (FISH) and RT-PCR analyses indicated a rearrangement of the MLL gene, but no fusion with previously identified MLL fusion partners at 17q, such as AF17 or MSF. Rapid amplification of cDNA ends (RACE) revealed an in-frame fusion of MLL to LASP1, a gene that is amplified and overexpressed in breast cancer. Retroviral transduction of myeloid progenitors demonstrated that MLL/LASP1 is the fourth known fusion of MLL with a cytoplasmic protein that has no in vitro transformation capability, thus establishing a potential subgroup among the MLL fusion proteins.     

Distinct gene expression profiling in chronic lymphocytic leukemia with 11q23 deletion.

Chronic lymphocytic leukemia (CLL) is a heterogeneous disease with regard to its clinical course. The limitations of the methods currently available for prognostic assessment in CLL do not allow accurate prediction of the risk of disease progression in individual patients. The recently developed cDNA array technique provides a unique opportunity to study gene expression in various malignancies. To identify new molecular markers for prognostication of CLL patients, we analyzed cDNA arrays by using hierarchical clustering and standard statistic t-test on 34 CLL patients. We found significant expression differences in 78 genes compared to the reference tonsillar B lymphocytes. A cluster of genes, LCP1, PARP, BLR1, DEK, NPM, MCL1, SLP76, STAM, HIVEP1, EVI2B, CD25, HTLF, HIVEP2, BCL2, MNDA, PBX3, EB12, TCF1, CGRP, CD14, ILB, GZMK, GPR17 and CD79B, was associated (P < 0.05) with the unfavorable 11q deletion and also with the unfavorable Binet stages B and C. We present here gene expression profiling that is associated with CLL patients with the 11q23 deletion. Many of the genes in the cluster have not previously been shown to be related to the initiation or progression of CLL. These novel findings provide fundamental information for further attempts to understand the interaction of the clustered genes in the leukomogenesis of CLL in order to better design treatments aimed at specific molecular target(s).     

Therapy-related adult acute lymphoblastic leukemia with t(4;11)(q21; q23): MLL rearrangement, p53 mutation and multilineage involvement.

A diagnosis of pro-B acute lymphoblastic leukemia (ALL) with CD15+ was made in a 42-year-old woman, 12 months after the treatment of uterine adenocarcinoma by carboplatinum, anthracyclines, etoposide and radiotherapy. Molecular cytogenetic studies revealed a karyotype with multiple chromosome changes, including the t(4;11)(q21;q23) and a 17p-chromosome, with MLL disruption and 17p13/p53 gene deletion in 86% of the cells. A p53 exon 6 mutation was documented, resulting in p53 protein stabilization, with 20% of the cells reacting with the 1801 anti-p53 monoclonal antibody. Dual-color FISH using MLL and p53 probes was performed on peripheral blood smears, providing direct evidence of the involvement of the blast cells and of the granulocytic lineage. Only a partial, shortlasting response was obtained by induction treatment, confirming that a poor prognosis is associated with therapy-related ALL with the 4;11 translocation.     

t(11;14)(q23;q24) generates an MLL-human gephyrin fusion gene along with a de facto truncated MLL in acute monoblastic leukemia

More than 20 different partner genes with MLL have been cloned from leukemia cells with translocations involving chromosome 11 band q23 (11q23). All reported partner genes fused in-frame to MLL and the fusion cDNA encode chimeric MLL proteins with a significant portion derived from the partner genes. We analyzed one patient with de novo acute monoblastic leukemia with t(11;14)(q23;q24) and identified that a human homologue of gephyrin (human gephyrin) fused with MLL. Gephyrin is a rat glycine receptor-associated protein, which forms submembranous complexes and anchor glycine or gamma-aminobutyric acidA receptors to microtubules. Alternative splicing of human gephyrin gene created two different forms of fusion cDNA. In one form, human gephyrin gene fused in-frame to MLL exon 9, and the chimeric product had COOH terminus of human gephyrin protein, including the tubulin binding site. In the other, the reading frame terminated shortly after the fusion point. As a result, only seven amino acids with no known function were attached to the NH2 terminus of MLL protein. The functional significance of this de facto truncated MLL gene product is not clear.